Advanced Behavioral Analyses Show that the Presence of Food Causes Subtle Changes in C. elegans Movement.

Angstman NB, Frank HG, Schmitz C - Front Behav Neurosci (2016)

Bottom Line:
In all seven investigated parameters, statistically significant differences were found in worm behavior between those moving on NGM-agar plates with an E. coli lawn and NGM-agar plates with no lawn.Furthermore, multiple test groups showed differences in interaction between variables as the parameters that significantly correlated statistically with speed of locomotion varied.The need to account for a nested design while performing statistical analyses in similar studies is also demonstrated.

Affiliation: Department of Neuroanatomy, Ludwig-Maximilians University of Munich Munich, Germany.

ABSTRACTAs a widely used and studied model organism, Caenorhabditis elegans worms offer the ability to investigate implications of behavioral change. Although, investigation of C. elegans behavioral traits has been shown, analysis is often narrowed down to measurements based off a single point, and thus cannot pick up on subtle behavioral and morphological changes. In the present study videos were captured of four different C. elegans strains grown in liquid cultures and transferred to NGM-agar plates with an E. coli lawn or with no lawn. Using an advanced software, WormLab, the full skeleton and outline of worms were tracked to determine whether the presence of food affects behavioral traits. In all seven investigated parameters, statistically significant differences were found in worm behavior between those moving on NGM-agar plates with an E. coli lawn and NGM-agar plates with no lawn. Furthermore, multiple test groups showed differences in interaction between variables as the parameters that significantly correlated statistically with speed of locomotion varied. In the present study, we demonstrate the validity of a model to analyze C. elegans behavior beyond simple speed of locomotion. The need to account for a nested design while performing statistical analyses in similar studies is also demonstrated. With extended analyses, C. elegans behavioral change can be investigated with greater sensitivity, which could have wide utility in fields such as, but not limited to, toxicology, drug discovery, and RNAi screening.

Figure 1: Representative images of C. elegans as analyzed in the present study. The panels show unprocessed frames of videos of wild type (N2) C. elegans on NGM-agar plates with an E. coli lawn (A) or with no lawn (B). The insets show individual worms (indicated by arrows) at higher magnification. The slightly uneven illumination of the panels stems from the fact that in the experimental setup of the present study, the field-of-view was placed offset from the middle of the agar plates. However, this had no impact on unequivocal identification and tracking of the worms on the plates (evaluation data not shown).

Mentions:
Plates were immediately placed under a dissecting microscope (MZ75; Leica, Wetzlar, Germany; equipped with 1.0X PlanApo objective) with an LCD light source with a color temperature of 2800 K (KL 1500; Schott, Mainz, Germany). Using the video capture function of the software, WormLab (Version 3.0.0, MBF Bioscience, Williston, VT, USA), 60 s long videos with a resolution of 1280 × 960 pixels were taken at 15 frames/s using a mono digital camera (Grasshopper 2, Point Grey Research, Richmond, BC, Canada). Using a horizontal mm ruler and the measure function of WormLab, videos were determined to have a scale of 8.37 μm/pixel, which enabled precise and accurate investigation of the parameters described in the next section. Accordingly, the field-of-view of the camera was 10.7 by 8.0 mm and, thus, 3% of the base area of the agar plates (Figure 1 and Supplementary Videos S1, S2).

Figure 1: Representative images of C. elegans as analyzed in the present study. The panels show unprocessed frames of videos of wild type (N2) C. elegans on NGM-agar plates with an E. coli lawn (A) or with no lawn (B). The insets show individual worms (indicated by arrows) at higher magnification. The slightly uneven illumination of the panels stems from the fact that in the experimental setup of the present study, the field-of-view was placed offset from the middle of the agar plates. However, this had no impact on unequivocal identification and tracking of the worms on the plates (evaluation data not shown).

Mentions:
Plates were immediately placed under a dissecting microscope (MZ75; Leica, Wetzlar, Germany; equipped with 1.0X PlanApo objective) with an LCD light source with a color temperature of 2800 K (KL 1500; Schott, Mainz, Germany). Using the video capture function of the software, WormLab (Version 3.0.0, MBF Bioscience, Williston, VT, USA), 60 s long videos with a resolution of 1280 × 960 pixels were taken at 15 frames/s using a mono digital camera (Grasshopper 2, Point Grey Research, Richmond, BC, Canada). Using a horizontal mm ruler and the measure function of WormLab, videos were determined to have a scale of 8.37 μm/pixel, which enabled precise and accurate investigation of the parameters described in the next section. Accordingly, the field-of-view of the camera was 10.7 by 8.0 mm and, thus, 3% of the base area of the agar plates (Figure 1 and Supplementary Videos S1, S2).

Bottom Line:
In all seven investigated parameters, statistically significant differences were found in worm behavior between those moving on NGM-agar plates with an E. coli lawn and NGM-agar plates with no lawn.Furthermore, multiple test groups showed differences in interaction between variables as the parameters that significantly correlated statistically with speed of locomotion varied.The need to account for a nested design while performing statistical analyses in similar studies is also demonstrated.

Affiliation:
Department of Neuroanatomy, Ludwig-Maximilians University of Munich Munich, Germany.

ABSTRACTAs a widely used and studied model organism, Caenorhabditis elegans worms offer the ability to investigate implications of behavioral change. Although, investigation of C. elegans behavioral traits has been shown, analysis is often narrowed down to measurements based off a single point, and thus cannot pick up on subtle behavioral and morphological changes. In the present study videos were captured of four different C. elegans strains grown in liquid cultures and transferred to NGM-agar plates with an E. coli lawn or with no lawn. Using an advanced software, WormLab, the full skeleton and outline of worms were tracked to determine whether the presence of food affects behavioral traits. In all seven investigated parameters, statistically significant differences were found in worm behavior between those moving on NGM-agar plates with an E. coli lawn and NGM-agar plates with no lawn. Furthermore, multiple test groups showed differences in interaction between variables as the parameters that significantly correlated statistically with speed of locomotion varied. In the present study, we demonstrate the validity of a model to analyze C. elegans behavior beyond simple speed of locomotion. The need to account for a nested design while performing statistical analyses in similar studies is also demonstrated. With extended analyses, C. elegans behavioral change can be investigated with greater sensitivity, which could have wide utility in fields such as, but not limited to, toxicology, drug discovery, and RNAi screening.